Modified Spectral Editing Methods for C CP/MAS Experiments in Solids

The spectral editing approach of Zilm and coworkers utilizes polarization, polarization inversion, and spin depolarization methods for enhancing or suppressing NMR spectral lines in solids. The proposed pulse sequences allow nonprotonated C, CH, CH₂, and CH₃ types of carbon resonances to be separated from one another and identified accordingly. The former method tentatively separates the nonprotonated C and CH₃ peaks with a cutoff shift of 35 ppm. This shift is a reasonable demarcation shift for a preponderance of organic molecules, but exceptions do exist that could constitute a serious drawback in a few instances. The new approach separates the nonprotonated C and CH₃ carbon peaks unequivocally using modified pulse sequences similar to those of Zilm. Further, both the CH only and CH₂ only spectra, respectively, can be acquired directly from combining so called (+) and (−) sequences using different spectral delay periods and pulse parameters. The (+) and the (−) pulse sequences produce signals for the nonprotonated and methyl carbons that have essentially the same amplitude but opposite phases. These spectra, combined with the previously reported CH₃ and nonprontonated C only spectra, offer a complete spectral editing technique for solid samples. Examples of these spectral editing methods are provided for 3-methylglutaric acid, fumaric acid monoethyl ester, and two complex natural products: methyl o-methylpodocarpate and 10-deacetylbaccatin III.     

Improvement of spectral editing in solids: A sequence for obtaining (13)CH + (13)CH(2)-only (13)C spectra

An improved spectral editing method for solids is described which allows one to obtain a set of subspectra in roughly two-thirds the amount of time as our original CPPI editing method for the same signal to noise. This improvement is afforded by a new pulse sequence that is used to acquire a (13)CH + (13)CH(2) spectrum which has very little (13)CH(3) or nonprotonated carbon contamination. By using this new sequence the (13)CH-only subspectrum is obtained much more efficiently. Criteria for optimizing the signal to noise in the edited subspectra are discussed. Copyright 2000 Academic Press.     

固体中13C CP/MAS谱编辑一些改进

本文将基本的谱编辑脉冲序列与消旋转边带的TOSS脉冲序列联用,可以实现强磁场下¹³C CP/MAS谱编辑.通常,固体中非质子化碳与甲基碳的自旋晶格弛豫时间差别很大,此差别可作为谱编辑过程中区分它们的一条附加判据.对具有复杂分子结构的体系,此判据的作用尤其重要.文中的实例将表明:一个完整的¹³C CP/MAS谱编辑一般可通过SCPTOSS谱(短接触时间交叉极化谱);SCPPITOSS (短接触时间交叉极化极化反转)谱,LCPDTOSS (长接触时间交叉极化及去极化)谱及部分弛豫的LCPDTOSS谱适当组合而得到.     

Improvement of Spectral Editing in Solids: A Sequence for Obtaining CH + CH2-Only C Spectra

An improved spectral editing method for solids is described which allows one to obtain a set of subspectra in roughly two-thirds the amount of time as our original CPPI editing method for the same signal to noise. This improvement is afforded by a new pulse sequence that is used to acquire a ¹³CH + ¹³CH₂ spectrum which has very little ¹³CH₃ or nonprotonated carbon contamination. By using this new sequence the ¹³CH-only subspectrum is obtained much more efficiently. Criteria for optimizing the signal to noise in the edited subspectra are discussed.     

Spectral editing in (13)C MAS NMR under moderately fast spinning conditions

Novel procedures for the spectral assignment of peaks in high-resolution solid-state (13)C NMR are discussed and demonstrated. These methods are based on the observation that at moderate and already widely available rates of magic-angle spinning (10--14 kHz MAS), CH and CH(2) moieties behave to a large extent as if they were effectively isolated from the surrounding proton reservoir. Dipolar-based analogs of editing techniques that are commonly used in liquid-state NMR such as APT and INEPT can then be derived, while avoiding the need for periods of homonuclear (1)H--(1)H multipulse decoupling. The resulting experiments end up being very simple, essentially tuning-free, and capable of establishing unambiguous distinctions among CH, CH(2), and --C--/-CH(3) carbon sites. The principles underlying such sequences were explored using both numerical calculations and experimental measurements, and once validated their editing applications were illustrated on a number of compounds.     

Accurate measurements of multiple-bond 13C-1H coupling constants from phase-sensitive 2D INEPT spectra.

Measurements of multiple-bond 13C-)H coupling constants are of great interest for the assignment of nonprotonated 13C resonances and the elucidation of molecular conformation in solution. Usually, the heteronuclear multiple-bond coupling constants were measured either by the J(CH) splittings mostly in selective 2D spectra or in 3D spectra, which are time consuming, or by the cross peak intensity analysis in 2D quantitative heteronuclear J correlation spectra (1994, G. Zhu, A. Renwick, and A. Bax, J. Magn. Reson. A 110, 257; 1994, A. Bax, G. W. Vuister, S. Grzesiek, F. Delaglio, A. C. Wang, R. Tschudin, and G. Zhu, Methods Enzymol. 239, 79.), which suffer from the accuracy problem caused by the signal-to-noise ratio and the nonpure absorptive peak patterns. Concerted incrementation of the duration for developing proton antiphase magnetization with respect to carbon-13 and the evolution time for proton chemical shift in different steps in a modified INEPT pulse sequence provides a new method for accurate measurements of heteronuclear multiple-bond coupling constants in a single 2D experiment.     

一种新的DEPT 谱编辑方法

提出一种新的基于多元线性回归计算的无畸变极化转移增益法(DEPT)谱编辑方法．该方法使用多元线性回归对谱图中的杂信号进行拟合,获得最佳拟合系数,然后将拟合系数代入拟合公式,通过该公式对DEPT 子谱进行计算,去除杂信号,最终分别得到纯净的CH₃、CH₂、CH 子谱图．实验结果证明,使用新方法对DEPT 谱进行编辑能够很好地减少杂信号的干扰,得到只含有CH_n 信号的子谱．新方法为碳谱的解析尤其是复杂和密集的谱线的解析提供了方便．     

Methylene spectral editing in solid-state 13C NMR by three-spin coherence selection

A robust new solid-state nuclear magnetic resonance (NMR) method for selecting CH2 signals in magic-angle spinning (MAS) 13C NMR spectra is presented. Heteronuclear dipolar evolution for a duration of 0.043 ms, under MREV-8 homonuclear proton decoupling, converts 13C magnetization of CH2 groups into two- and three-spin coherences. The CH2 selection in the SIJ (C H H) spin system is based on the three-spin coherence S(x)I(z)J(z), which is distinguished from 13C magnetization (S(x)) by a 1H 0 degrees/90 degrees pulse consisting of two 45 degrees pulses. The two-spin coherences of the type S(y)I(z) are removed by a 13C 90 degrees x-pulse. The three-spin coherence is reconverted into magnetization during the remainder of the rotation period, still under MREV-8 decoupling. The required elimination of 13C chemical-shift precession is achieved by a prefocusing 180 degrees pulse bracketed by two rotation periods. The selection of the desired three-spin coherence has an efficiency of 13% theoretically and of 8% experimentally relative to the standard CP/MAS spectrum. However, long-range couplings also produce some three-spin coherences of methine (CH) carbons. Therefore, the length of the 13C pulse flipping the two-spin coherences is increased by 12% to slightly invert the CH signals arising from two-spin coherences and thus cancel the signal from long-range three-spin coherences. The signal intensity in this cleaner spectrum is 6% relative to the regular CP/TOSS spectrum. The only residual signal is from methyl groups, which are suppressed at least sixfold relative to the CH2 peaks. The experiment is demonstrated on cholesteryl acetate and applied to two humic acids.     

HAT HMBC: a hybrid of H2BC and HMBC overcoming shortcomings of both

A new 2D NMR experiment, HAT HMBC, that is a hybrid of H2BC and HMBC aims at establishing two-bond correlations absent in H2BC spectra because of vanishing (3)J(HH) coupling constants. The basic idea is to create an additional pi phase difference in the multiplet structure in HMBC peaks with respect to the (n+1)J(HH) coupling constant between the proton(s) attached to a (13)C and a (1)H separated by n bonds. Thus HMBC peaks associated with small J(HH) will be the most attenuated in a HAT HMBC spectrum in comparison to a regular HMBC spectrum, i.e. peaks associated with (n+1)J(HH) and (n)J(CH) will for n>2 usually be strongly attenuated. The HAT HMBC pulse sequences contain the same number of pulses as regular HMBC and are only a few milliseconds longer.     

Experimental access to HSQC spectra decoupled in all frequency dimensions

A new operator called RESET “Reducing nuclEar Spin multiplicitiEs to singuleTs” is presented to acquire broadband proton decoupled proton spectra in one and two dimensions. Basically, the homonuclear decoupling is achieved through the application of bilinear rotation pulses and delays. A [BIRD]^r,x pulse building block is used to selectively invert all proton magnetization remotely attached to ¹³C isotopes, which is equivalent to a scalar J decoupling of the protons directly attached to ¹³C from all other protons in the spin system. In conjunction with an appropriate data processing technique pure shift proton spectra are obtained. For this purpose, the concept of constant time acquisition in the observe dimension is exploited. Both ideas were merged together producing superior HSQC based pseudo 3D pulse sequences. The resulting HSQC spectra show cross peaks with collapsed multiplet structures and singlet responses for the proton chemical shift frequencies. An unambiguous assignment of signals from overcrowded spectra becomes much easier. Finally, the recently introduced SHARC technique is exploited to enhance the capability of the scalar J decoupling method. A significant reduction of the total measurement time is achieved. The time is saved by reducing the number of ¹³C chemical shift evolution increments and working with superimposed narrow spectral bandwidths in the ¹³C indirect domain.     

最高量子相关谱技术在同核偶合常数测定中的应用(英文)

偶合常数是一个重要的NMR参数,其数值与分子中化学键的二面角有关,可以为分子结构研究提供很重要的信息.多维NMR谱由于具有较大的化学位移分辨率,因此常常被用来测定同核或异核自旋-自旋偶合常数.本文介绍了利用最高量子相关技术(MAXY)测定同核偶合常数的方法.MAXY是最近发展的一种多维NMR谱编辑技术,可以使不同官能团(CH,CH₂,CH₃)的相关峰分布于不同的图谱区域,因此比常规的二维谱具有更高的化学位移分辨率.而且被分离开来的NMR相关峰呈吸收性线型,能清楚地展示各自的偶合分裂特征,可以直接用于测定偶合常数.     

Determination and assignment of heteronuclear long-range coupling constants. Methods based on semiselective INEPT

One-dimensional methods for the determination and assignment of heteronuclear ¹H-X (X = rare spin-z) coupling constants based on the semiselective polarization transfer via INEPT pulse sequence are proposed. Here the selectivity of the polarization transfer plays a positive role with respect to the sensitivity of the measurement and purity of the observed multiplets. In nonrefocused experiments the acquired antiphase multiplets enable an unambiguous assignment of long-range couplings of a preselected proton. The analysis of such multiplets is also discussed. In the refocused version the purging pulse (INEPT+) was used to provide pure in-phase multiplets. The spectral editing technique DISCO was applied to simplify the spectra and to extract the couplings from complex multiplets. Finally, the modified INEPT experiments which combine semiselective polarization transfer with selective proton decoupling are proposed.     

Incorporating homonuclear polarization transfer into PRESS for proton spectral editing: illustration with lactate and glutathione

A proton spectral editing pulse sequence for the detection of metabolites with spin systems that involve weak coupling is presented. The sequence is based on homonuclear polarization transfer incorporated into the standard PRESS (Point RESolved Spectroscopy) sequence, which is a volume-selective double spin echo method, to enable spatial localization. All peaks in the region of interest are initially suppressed whether they are peaks from the target metabolite or from contaminating background. The target signal is then restored by polarization transfer from a proton that has a resonance outside the suppressed region and to which the target spins are weakly coupled. This is achieved by the application of a 90 degrees hard pulse with phase orthogonal to that of the PRESS excitation pulse at the location of the first echo in PRESS and by optimizing the two PRESS timings, TE(1) and TE(2), for most efficient yield. Background signal not coupled to any protons outside the initially saturated region remains suppressed. The advantage of this sequence compared to multiple quantum filters is that signal from singlet peaks outside the suppressed area are preserved and can thus be used as a reference. The efficacy of the sequence was verified experimentally on phantom solutions of lactate and glutathione at 3.0 T. For the AX(3) spin system of lactate, the sequence timings were optimized by product operator calculations whereas for the ABX spin system of the cysteinyl group of glutathione numerical calculations were performed for sequence timing optimization.     

Spectral Editing in C MAS NMR under Moderately Fast Spinning Conditions

Novel procedures for the spectral assignment of peaks in high-resolution solid-state ¹³C NMR are discussed and demonstrated. These methods are based on the observation that at moderate and already widely available rates of magic-angle spinning (10–14 kHz MAS), CH and CH₂ moieties behave to a large extent as if they were effectively isolated from the surrounding proton reservoir. Dipolar-based analogs of editing techniques that are commonly used in liquid-state NMR such as APT and INEPT can then be derived, while avoiding the need for periods of homonuclear ¹H–¹H multipulse decoupling. The resulting experiments end up being very simple, essentially tuning-free, and capable of establishing unambiguous distinctions among CH, CH₂, and carbon sites. The principles underlying such sequences were explored using both numerical calculations and experimental measurements, and once validated their editing applications were illustrated on a number of compounds.     

A phase-insensitive single-scan method for volume-selective editing of NMR signals using cyclic polarization transfer. In vivo determination of lactate

A phase-insensitive pulse sequence serving the volume-selective editing of resonances in biomedical NMR spectroscopy is described. The editing principle is the temporary transfer of polarization of the spins to be detected to other species coupled to them. The technique is of particular interest for the localized detection of the lactate methyl proton line while spoiling coherences of uncoupled as well as coupled lipid nuclei otherwise overlapping the lactate resonance. The whole line-editing and coherence-spoiling procedure takes place completely in each scan of any accumulation or phase-cycling series. Phantom studies were carried out with an experimental 4.7 T system. The technique was also tested with volunteers in whole-body tomographs operating at 2 and 3 T. Comparative spectra are presented. It is concluded that editing is crucial for the reliable determination of lactate in muscle and in brain.     

5- and 6-pulse electron spin echo envelope modulation (ESEEM) of multi-nuclear spin systems

In 3-pulse ESEEM and the original 4-pulse HYSCORE, nuclei with large modulation depth (k approximately 1) suppress spectral peaks from nuclei with weak modulations (k approximately 0). This cross suppression can impede the detection of the latter nuclei, which are often the ones of interest. We show that two extended pulse sequences, 5-pulse ESEEM and 6-pulse HYSCORE, can be used as experimental alternatives that suffer less strongly from the cross suppression and allow to recover signals of k approximately 0 nuclei in the presence of k approximately 1 nuclei. In the extended sequences, modulations from k approximately 0 nuclei are strongly enhanced. In addition, multi-quantum transitions are absent which simplifies the spectra. General analytical expressions for the modulation signals in these sequences are derived and discussed. Numerical simulations and experimental spectra that demonstrate the higher sensitivity of the extended pulse sequences are presented.     

选择性1NEPT技术及其在FX—90Q仪器上的应用

本文介绍一种~1H—~(13)C远程极化转移的脉冲序列.在JEOL FX—90QNMR波谱仪上实现了这种技术.对一些典型化合物的测定表明,选择性非灵敏核的远程极化转移增强法(SEL1NEPT)在测定季碳原子和进行谱带归属中,是一个有力的工具.     

Multiple-rotor-cycle 2D PASS experiments with applications to (207)Pb NMR spectroscopy

Thetwo-dimensional phase-adjusted spinning sidebands (2D PASS) experiment is a useful technique for simplifying magic-angle spinning (MAS) NMR spectra that contain overlapping or complicated spinning sideband manifolds. The pulse sequence separates spinning sidebands by their order in a two-dimensional experiment. The result is an isotropic/anisotropic correlation experiment, in which a sheared projection of the 2D spectrum effectively yields an isotropic spectrum with no sidebands. The original 2D PASS experiment works best at lower MAS speeds (1-5 kHz). At higher spinning speeds (8-12 kHz) the experiment requires higher RF power levels so that the pulses do not overlap. In the case of nuclei such as (207)Pb, a large chemical shift anisotropy often yields too many spinning sidebands to be handled by a reasonable 2D PASS experiment unless higher spinning speeds are used. Performing the experiment at these speeds requires fewer 2D rows and a correspondingly shorter experimental time. Therefore, we have implemented PASS pulse sequences that occupy multiple MAS rotor cycles, thereby avoiding pulse overlap. These multiple-rotor-cycle 2D PASS sequences are intended for use in high-speed MAS situations such as those required by (207)Pb. A version of the multiple-rotor-cycle 2D PASS sequence that uses composite pulses to suppress spectral artifacts is also presented. These sequences are demonstrated on (207)Pb test samples, including lead zirconate, a perovskite-phase compound that is representative of a large class of interesting materials.     

Selective Excitation of Protons Directly Bonded to Carbon-13

New pulse sequences for the selective excitation and editing of NMR spectra of protons directly bonded to carbon-13 in CH, CH₂, and CH₃ groups are described. Experimental results as well as theoretical calculations demonstrate that these sequences have excitation profiles of about ±0.5J_CH or less and require no pulse shaping or special hardware to implement. After the isotope-directed excitation of the protons has been generated, the proton magnetization may be relayed to other protons via an isotropic mixing sequence to yield 1D HOHAHA subspectra or 2D COSY subspectra. Examples of applications to compounds containing carbon-13 either at natural abundance or with enrichment are shown.     

A novel dipolar dephasing method for the slow magic angle turning experiment

Complete suppression of the resonances from protonated carbons in a slow magic angle spinning experiment can be achieved using five dipolar dephasing (Five-DD) periods distributed in one rotor period. This produces a spectrum containing only the spinning sidebands (SSB) from the nonprotonated carbons. It is shown that the SSB patterns corresponding to the nonprotonated carbons are not distorted over a wide range of dipolar dephasing times. Hence, this method can be used to obtain reliable principal values of the chemical shift tensors for each nonprotonated carbon. The Five-DD method can be readily incorporated into isotropic-anisotropic 2D experiments such as FIREMAT and 2D-PASS to facilitate the measurement of the (13)C chemical shift tensors in complex systems.